The major objective of this research proposal is to determine the biochemical characteristics and covalent structure of two genetically distinct collagenous polypetide C and D chains from normal glomerular basement membrane. The diabetic nephrophatic basement membrane collagen chains will be examined for alterations in the amino acid sequence, changes in hydroxylation of proline and lysine residues, glycosylation of hydroxylysine, and the ratio of C and D chains. The C and D chains will be prepared by pepsin digestion of glomerular basement membrane and corteces. The purified chains will be cleaved with CNBr and the CNBr peptides will be isolated and characterized. The CNBr peptides of C and D chain prepared from normal and diabetic basement membrane collagens will be analyzed and compared for the hydroxylation of prolyl and lysyl residues, glycosylation of hydroxylysine and for the alteration in the primary structure. The glomeruli preparation from normal and diabetic kidneys will be analyzed for the nature and the extent of lysine and hydroxylysine derived cross-links in the basement membrane. Biosynthetic studies in tissue culture using viable glomeruli prepared from normal and naturally occurring diabetic rats will be performed to determine the rate of synthesis of C and D chains, extent of hydroxylation of proline and lysine residues and the glycosylation of hydroxylysine in normal and diseased state. The nature and extent of lysine and hydroxylysine derived crosslinks formed during biosynthesis will be also examined.